Skin is one of the largest organs in the body and covers substantially the entire body surface. Skin is composed of two main layers: the surface epithelium or epidermis, which contains keratinocytes as one type of epidermal cells, and the subjacent connective tissue layer or dermis, which contains fibroblasts as one type of dermal cells. The functions of skin include protecting an organism from injury and dessication by serving as a barrier to infection, perceiving or detecting environmental stimuli, excreting various substances, regulating body temperature, and helping to maintain water balance. Because of its quantitative and qualitative importance, substantially intact and healthy skin is crucial, not only for the well being of an organism but for its very survival.
The health and integrity of skin may be compromised by congenital or acquired pathological conditions, either acute or chronic, for which normal skin regeneration and repair processes may be inadequate. These conditions include, but are not limited to, burns, wounds, ulcers, infections, and/or congenital abnormalities. Patients who are burned over a large surface area often require immediate and extensive skin replacement. Less life-threatening but chronic skin conditions, as occur in venous stasis ulcers, diabetic ulcers, or decubitus ulcers as three examples, may progress to more severe conditions if left untreated, particularly since patients with these conditions have an underlying pathology. Reduction of morbidity and mortality in such patients depends upon timely and effective restoration of the structure and function of skin.
Skin substitutes derived either ex vivo or in vitro may be used to treat these or other conditions. Desirable skin substitutes are readily available, require a minimum amount of donor skin, are otherwise simple to produce, and are cost-effective to generate and maintain. Several approaches to produce skin substitutes which satisfy some or all of these requirements have been attempted, with varying degrees of success. However, no skin substitute has yet regenerated all of the structures and functions of skin. Rather, all are subsets of uninjured skin. Only a transplant of full thickness skin, which demonstrates scarring during healing, and restores virtually all structures and functions of normal uninjured skin.
Materials have been manufactured for use in permanent skin repair. These materials contain different components replacing or simulating the components and functions of the dermis and/or epidermis. Examples of these materials include EpiCel™, which lacks a dermal component and uses the patient's own cultured keratinocytes; Integra™, which uses a collagen-glycosaminoglycan (GAG) matrix to provide an acellular dermal component and uses a thin epidermal autograft; AlloDerm™, which uses a dermal matrix and a thin epidermal autograft; DermaGraft™, which uses a polyglycolic acid/polylactic acid (PGA/PLA) matrix and allogeneic human fibroblasts for the dermis; Hyaff/LaserSkin™, which uses hyaluran and fibroblasts for the dermis, and hyaluran and the patient's own keratinocytes for the epidermis; and PolyActive™, which uses polyethylene oxide/polybutylthalate (PEO/PBT) and the patient's own fibroblasts for the dermis, and the patient's cultured keratinocytes for the epidermis.
Materials to either temporarily cover wounds, or to stimulate permanent skin repair processes, included ApliGraft™, which uses collagen gel and allogeneic fibroblasts for the dermis, and cultured allogeneic keratinocytes for the epidermis; Comp Cult Skin™ or OrCel™, which uses collagen and allogeneic fibroblasts for the dermis, and cultured allogeneic keratinocytes for the epidermis; and TransCyte™, which uses allogeneic fibroblasts for the dermis and a synthetic material, BioBrane™, for the epidermis.
While the above materials are useful to varying degrees, each has disadvantages and limitations. Some of the materials are mechanically fragile, making it difficult to perform the required manipulations and transfers of the material in large sections without tearing. Instead, the materials must be used as smaller pieces, which makes coverage of large surface areas technically laborious for the physician and cosmetically undesirable for the patient. The materials are also susceptible to microbial contamination, which is unacceptable for patients who are already at an increased risk for infection due to their compromised condition. The materials show varying rates of engraftment and time to heal, both of which must be considered in balancing the advantage of a particular material over another for a particular patient. For example, a material which is otherwise acceptable but which takes longer to engraft and heal is less desirable, since recovery includes as rapid a return to a normal routine as possible.
Yannas et al. in U.S. Pat. No. 4,458,678 discloses a method for preparing a fibrous lattice and seeding it with viable cells. The lattice is prepared by pouring an aqueous slurry of collagen and glycosaminoglycan into an open metal tray or pan. Use of an open tray results in asymmetric contact of the slurry to ambient conditions; that is, one surface of the slurry is in contact with a surface of the tray or pan (“pan surface”), while another surface of the slurry is in contact with the atmospheric environment (“air surface”). The pan is then placed in contact with a heat sink, such as the refrigerated shelf of a lyophilizer, to initiate freezing of the aqueous slurry from the pan side. Freezing proceeds predominantly in a vectorial direction from the pan surface to the air surface, with heat flowing much more rapidly toward the pan surface than the air surface. Upon freezing, the structure of the lattice is asymmetric with, generally, more frequently distributed reticulations of polymers toward the pan surface than the air surface. After freeze-drying, the resulting lattice is an asymmetric structure, which results in non-uniform ability for cells to penetrate the lattice. Furthermore, the method and apparatus of Yannas limit both the uniformity of thickness, and the minimum thickness that can be obtained consistently and deliberately.
The inventor's own previous composite skin replacement, disclosed in U.S. Pat. No. 5,976,878, which is expressly incorporated by reference herein in its entirety, has been used successfully for permanent skin replacement. It is applied surgically in a single procedure, and contains a layer of cultured epidermal cells, a synthetic dermal membrane component, and a substantially nonporous synthetic lamination layer on one surface of the dermal membrane component. The synthetic dermal membrane component is formed from collagen, or collagen and a mucopolysaccharide compound, and is laminated with the same collagen or collagen and mucopolysaccharide compound-containing solution containing a volatile cryoprotectant. The substantially nonporous lamination layer may be located between the dermal component and the layer of cultured epidermal cells, promoting localization of epidermal cells on the surface of the dermal component and movement of nutrients to the cells of the cellular epidermal component. This composition can also be used to deliver biologically active molecules to the site where it is applied. The apparatus for preparing the synthetic dermal membrane component, used to prepare the composite skin replacement of the '878 patent, is disclosed in U.S. Pat. No. 5,711,172, which is expressly incorporated herein by reference in its entirety.
Desirable features of the above-described composite skin replacement included an increased rate of vascularization of the area covered by the material, decreased microbial contamination, increased nutrient supply, and improved epidermal barrier function, compared to other materials. Areas covered with the composite skin replacement required less time to engraft and heal, and the material was less susceptible to microbial contamination compared to other materials. Other desirable features were that this material was relatively non-fragile and easy to handle, and could be generated relatively rapidly, for example, within the time frame in which a burn patient would require skin grafts. However, while no other alternative material has healed excised, full thickness wounds more rapidly, and with as low an incidence of microbial contamination, limitations still exist. Thus, there remains a need to more closely approach the structural and functional properties of normal uninjured skin.